白粉病菌诱导的小麦抑制差减文库构建与杂交点阵膜制备

A SSH cDNA Library Construction and Array Preparation with Wheat Challenged by Erysiphe graminis DC

以本实验室培育的抗白粉病品系“百农 32 17×Mardler”BC5F4为材料 ,构建了一个白粉病菌诱导的抑制差减杂交cDNA文库。建库过程质量分析表明 ,差减杂交效率较高 ,文库质量较好。文库滴度为 4 .2× 10 9cfu·ml-1;插入片段平均为 30 0bp左右。挑取文库阳性克隆 96 0个 ,扩增插入片段 ,将其制备成高密度杂交点阵膜。通过探针杂交检测表明 ,本研究制备点阵膜的方法简便可靠 。

A suppression subtractive hybridization (SSH) cDNA library was constructed with a powdery mildew resistance line: 'Bai Nong 3217×Mardler' BC 5F 4 wheat leaf challenged by Erysiphe graminis DC at the primary stage. The process of the library construction was checked and analyzed. The results showed that the technique was efficient in the subtractive hybridization. The titer of the SSH cDNA library was 4.2×10 9 cfu·ml -1 , and the average insert size was around 300 bp. About 960 positive clones were picked, and the PCR products of their inserts were used to make array membranes. After checking the array hybridized with the probe, the method that was set up to make the array was credible and relative simple, and the array membranes can be used to screen disease resistance genes further.